Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:5.99.1.2 (topoisomerase)
9,166 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Structures retaining many of the morphological features of nuclei may be released by lysing human cells in a non-ionic detergent and 2 M NaCl. Such nucleoids contain all the nuclear DNA packaged within a flexible cage of RNA and protein. HeLa nucleoids have been spread at an air-water interface and viewed in the electron microscope. A tangled network of superhelical fibres surrounds the collapsed cage. Irradiation with gamma-rays abolishes supercoiling and treatment with the untwisting enzyme or a low concentration of ethidium reduces it. A high concentration of ethidium induces supertwisting. The nuclear DNA of higher cells can be isolated naked, supercoiled and intact.
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PMID:Electron-microscopy of intact nuclear DNA from human cells. 23 Oct 42

Topotecan (SK&F 104864), a water-soluble analogue of the topoisomerase I inhibitor camptothecin, is currently in Phase II clinical trial for solid tumors. We have characterized topotecan in terms of its effect upon gamma-radiation-induced cell killing. In colony formation experiments, subtoxic concentrations of topotecan (2 microM) potentiated radiation-induced killing of exponentially growing Chinese hamster ovary or P388 murine leukemia cultured cells. Survival curve shoulders were reduced; the slopes of the exponential portions of the curves were decreased to a small extent. D37 and D10 (radiation dose resulting in 37 and 10% survival of colony-forming ability) values were reduced by approximately 60 and 50%, respectively, in the case of Chinese hamster ovary cells. In P388 cells, topotecan reduced D37 by 35 to 40% and D10 by 20 to 25%. Potentiation of radiation-induced cell killing by topotecan was absolutely dependent upon the presence of the topoisomerase I inhibitor during the first few (less than 30) min after irradiation. Association of topoisomerase I with this effect was confirmed in studies of Chinese hamster ovary cells previously made resistant to camptothecin (and cross-resistant to topotecan), resulting in decreased cellular content of topoisomerase I. These cells were found to be 2- to 3-fold hypersensitive to gamma-radiation-induced killing. P388 camptothecin-resistant cells were further sensitized to the lethal effects of ionizing radiation by nontoxic treatment with the topoisomerase II inhibitor novobiocin, consistent with increased dependence of topoisomerase I-deficient cells upon topoisomerase II.
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PMID:Synergistic cell killing by ionizing radiation and topoisomerase I inhibitor topotecan (SK&F 104864). 165 71

Changes in linking number and the apparent winding angle of pBR322 DNA have been evaluated in mixed ethanol-water solvents containing either Na or Mg as the major counterion contributing to the electrostatic shielding of the duplex. The average number of superhelical turns (tau) produced in the standard electrophoresis buffer (Tris-borate-EDTA, pH 8.0) by the transfer of DNA, relaxed in 200 mM NaCl, 10 mM NaH2PO4/Na2HPO4, and 2 mM EDTA, pH 7, by calf thymus topoisomerase or ligated in 6.6 mM MgCl2, 1 mM KCl, 1 mM ATP, 1 mM dithiothreitol, and 66 mM Tris, pH 7.6, by T4 ligase, was determined as a function of the EtOH concentration. At low enzyme concentrations, the tau values became increasingly more positive in the presence of both cations as the ethanol concentration increased, indicating that the duplex structure was overwound in the ethanol solvents. Winding angle changes between 0 and 20% ethanol, calculated from these values of tau, exhibited the same correlations with CD spectral properties as had been previously observed for 100% aqueous systems containing monovalent cations [Kilkuskie, R., Wood, N., Shinn, R., Ringquist, S., & Hanlon, S. (1988) Biochemistry 27, 4377-4386]. The results at higher concentrations of ethanol (25-30%), however, were anomalous for the Mg-ligase system. The anomalies increased with higher ethanol, ligase, or Mg concentration. Gel run under these conditions showed enhanced concentrations of slow-moving components, indicative of ligation of intermolecular associated DNA species. At a 10-fold higher level of ligase, ethanol appeared to unwind the duplex, confirming the results of Lee, Mizusawa, and Kakefuda [(1981) Proc. Natl. Acad. Sci. U.S.A. 78, 2838-2842]. All of these anomalies occur under solvent conditions which are close to conditions which produce a heterogeneous dispersion of sedimenting species in ultracentrifugal experiments and compact rodlike structures, visualized by electron microscopy. The circular dichroism spectra at the onset of the formation of these structures show the characteristics of a chirally packed array of DNA duplexes. The reversal of the trend of the ethanol effect on linking number at higher enzyme and Mg(II) concentrations can be most easily explained by the promotion of the condensation phenomenon by either the ligase or a contaminating factor in the preparation. We suggest that the anomalies in the linking number and winding angle values are due to either ligation of chirally bent DNA species or a change in the helical period as the linear DNA adapts to the conformation required for collapse.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Linking number anomalies in DNA under conditions close to condensation. 265 31

A preparation of bacteriophage T4-induced deoxyribonucleotide synthetase complex is described. This very large complex of enzymes can be separated by centrifugation at 100,000 X g, by sucrose step gradient centrifugation, or with molecular exclusion columns. By direct assay and by unidimensional and two-dimensional acrylamide electrophoretic separations the following T4-coded enzymes were shown to be associated with the complex: ribonucleoside diphosphate reductase, dCMP deaminase, dCTP/dUTPase, dCMP hydroxymethylase, dTMP synthetase, and DNA polymerase. Other phage-coded prereplicative proteins related to DNA replication and other phage functions such as the proteins coded by genes 32, 46, rIIA, and rIIB as well as many unidentified proteins were also consistently associated with the isolated fractions. T4 DNA topoisomerase, a membrane-bound enzyme, was found in quantity in all purified fractions of the complex, even in preparations apparently free of membrane and of T4 DNA. The functional integrity of a segment of the complex was followed by measuring the conversion of [5-3H]CDP to the level of 5-hydroxymethyl dCMP. This series of reactions requires the actions of T4-coded ribonucleoside diphosphate reductase and its associated reducing system, dCTP/dUTPase and dCMP hydroxymethylase, 3H being lost to water at the last step. In this reaction sequence an intermediate, [5-3H]dCMP, is maintained at low steady state concentrations, and argument is presented that the synthesis of deoxyribonucleotides is channeled and normally tightly coupled to DNA replication. One of the primary characteristics of this complex is its ready dissociation of dilution into smaller complexes of proteins and to the free forms of the proteins. That the complex is held together by weak electrostatic forces was supported by its sensitivity to dissociation at moderate salt concentrations. Not only the enzymes required in deoxyribonucleotide synthesis but T4 DNA polymerase, T4 DNA topoisomerase, and a number of other proteins dissociate to varying degrees from the larger complexes under these conditions.
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PMID:Characteristics of a bacteriophage T4-induced complex synthesizing deoxyribonucleotides. 675 52

A water extract of Coptis chinensis was found to have the ability to stabilize the cleavable complex with mammalian DNA topoisomerase I. As the result of bioassay-guided fractionation, two protoberberine alkaloids, epiberberine and groenlandicine, were identified as active principles with topoisomerase I-mediated DNA cleavage activity in vitro. These two alkaloids did not induce topoisomerase II-mediated DNA cleavage. During further examination of the structurally related protoberberine alkaloids, berberrubine which is produced during the processing of Coptis rhizome as traditional medicine, was identified as a specific inducer of topoisomerase II-mediated DNA cleavage in vitro. These results indicated that protoberberine alkaloids are a chemical family which can induce cleavable complexes with topoisomerases I and II.
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PMID:Inhibitors of DNA topoisomerase I and II isolated from the Coptis rhizomes. 748 Feb 1

Electron-affinic compounds with strong DNA intercalating properties have demonstrated less than the expected radiosensitization due to restriction of their mobility along the DNA backbone and their lower extravascular diffusion in tumors. A 2-nitroimidazole linked 1,2,3,4-tetrahydroacridine derivative (THNLA-1) has been synthesized as a hypoxia-selective cytotoxin and radiosensitizer with presumably lower DNA-binding affinity due to the perturbation of the planarity in the acridine ring. THNLA-1 is a good hypoxia-selective cytotoxin with a differential toxicity of approximately equal to 11 in V79 cells, but it is approximately equal to 2 times less potent on a concentration basis than NLA-1 (the 2-nitroimidazole linked acridine analog). However, THNLA-1 is a very efficient radiosensitizer, showing a sensitization enhancement ratio (SER) of 3.04 +/- 0.05 at 100 microM at 25 degrees C, and the concentration giving an SER of 1.6(C1.6) is 19.0 +/- 0.5 microM. The therapeutic index, defined as the ratio of the clonogenic IC50 under aerobic conditions for 1-h exposure (IC50A,1h) to the C1.6 value, is 20 for THNLA-1 vs. 11 for NLA-1. THNLA-1's partition coefficient in octanol/water is 0.14 +/- 0.02. Topoisomerase I and II interaction studies with THNLA-1 showed that topoisomerase I-mediated relaxation of supercoiled DNA was inhibited at relatively high THNLA-1 concentrations (> or = 1000 microM), while topoisomerase II-mediated decatenation of kinetoplast DNA remained unaffected even in concentrations toxic in vitro under aerobic conditions. Uptake studies under aerobic conditions showed high intracellular drug concentrations, compatible with the required ones for topoisomerase I inhibition.
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PMID:9-[3-(2-Nitro-1-imidazolyl)propylamino]-1,2,3,4-tetrahydroacridine hydrochloride. A novel DNA-affinic hypoxic cell cytotoxin and radiosensitizer. Comparison with NLA-1. 770 30

A series of analogs based on a novel template, 11-aza-(20S)-camptothecin, were obtained from total synthesis and tested as potential anticancer drugs in the topoisomerase I enzyme cleavable complex assay. The parent compound 11-aza-(20S)-camptothecin (8) was derived from a Friedlander condensation between the known aminopyridine derivative 3-(3-amino-4-picolylidene)-p-toluidine and optically active tricyclic ketone 7. Compound 8 had activity approximately twice that of (20S)-camptothecin in the calf thymus topoisomerase I cleavable complex assay. Compounds were prepared wherein the 11-aza nitrogen atom was quaternized as either the corresponding N-oxide or methyl iodide. Compounds with quaternized N-11 showed improved water solubility and were equipotent to the clinically investigated camptothecin analog topotecan in the cleavable complex assay. These compounds were evaluated in vivo in nude mice bearing HT-29 human colon carcinoma xenografts. The analog 11-aza-(20S)-camptothecin 11-N-oxide was found to significantly retard tumor growth when compared to untreated controls. Finally, 7,10-disubstituted 11-azacamptothecin analogs were synthesized using Pd(0) coupling reactions of 10-bromo-7-alkyl-11-aza-(20S)-camptothecins 19 and 20, which in turn were available from a Friedlander condensation of the novel bromopyridine derivatives 17a and 17b with 7. Among the 10-substituted series, a number of analogs displayed extremely high in vitro potency against topoisomerase I and improved aqueous solubility. A significant number of the compounds were found to be active in whole cell cytotoxicity assays and several were evaluated in nude mice bearing the HT-29 tumor xenografts. The most effective of these proved to be (S)-11-aza-7-ethyl-10-(aminohydroximinomethyl)camptothecin trifluoracetic acid salt (27), a potent topoisomerase I inhibitor which demonstrated excellent efficacy in both short term and in extended in vivo assays. A comparison between in vitro enzyme data and in vivo data from nude mouse studies in other compounds in this series revealed a poor overall correlation between topoisomerase inhibition in vitro and antitumor efficacy in vivo.
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PMID:Synthesis, topoisomerase I inhibitory activity, and in vivo evaluation of 11-azacamptothecin analogs. 770 14

Resistance to chemotherapy in brain tumors is complex and may involve multiple mechanisms. For commonly used drugs, such as nitrosoureas and platinum compounds, major mechanisms may involve increaded DNA repair or removal of the drug-DNA adducts. For water soluble nitrosoureas and also for platinum compounds, other mechanisms, such as alteration in drug transport, may be important. Another major mechanism may involve glutathione and glutathione-S-transferase pathways. For vinca alkaloids and epipodophyllotoxins p-glycoprotein mediated MDR appears to be the major feature in drug resistance. In addition, alteration of tubulin and topoisomerase II have been described in resistance to vinca alkaloids and epipodophyllotoxins respectively. Recently, increased multidrug resistance associated protein gene expression has been found in glioma cells and brain tumor samples; its clinical significance requires further investigation.
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PMID:Drug resistance in brain tumors. 780 93

The topoisomerase I inhibitor topotecan is a potent water-soluble camptothecin derivative with activity in a wide variety of preclinical models. Topotecan exhibits schedule dependency in vivo, with the greatest activity being observed on repeated dose schedules. On the basis of the initial clinical studies that showed a short plasma half-life, we attempted to prolong drug exposure by giving topotecan as a 24-h infusion weekly. In a phase I trial, we treated 32 patients at doses ranging from 1.0 to 2.0 mg/m2. The patient population had not been heavily pretreated with chemotherapy and was of good performance status. The incidence of neutropenia, which was dose-limiting, increased sharply with relatively small increments in dose. Doses greater than 1.5 mg/m2 were associated with nadirs that developed after one to three weekly treatments. A patient with metastatic colorectal cancer had a prolonged partial response. The plasma pharmacokinetics of topotecan (lactone and open forms) was characterized in 21 patients. Mean plasma steady-state drug levels were proportional to the dose and were within the range required to exert cytotoxicity in preclinical models. Plasma elimination curves were fit to a one-compartment model, in which the harmonic mean half-life of topotecan was 3.5 h. The ratio of the lactone to the total drug concentrations was constant throughout, which suggests that for this schedule the total drug concentration may be used as a measure of active lactone exposure. This conclusion is supported by the pharmacodynamic analysis, which revealed a positive correlation of both lactone and total drug steady-state concentrations with bone marrow toxicity. The further investigation of this and other infusional schedules in phase II trials will be conducted. The steady-state concentrations of total drug will be measured in several of these trials to establish its potential role in adaptive dosing using this schedule. Such a strategy is justified by the interpatient variability in toxicity and the steep dose-response curve observed in this study. Preliminary evidence of interpatient variability in the mRNA expression of topoisomerase I in the peripheral mononuclear cells and colon mucosa is presented. Trials are under way using biological endpoints for further selection of patients in whom the use of topoisomerase inhibitors may be therapeutically beneficial.
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PMID:Clinical, pharmacokinetic and biological studies of topotecan. 807 27

Topoisomerase I and topoisomerase II allow a metabolically active cell to mobilize its supercoiled chromosomal DNA and undergo replication, transcription, recombination, and repair. Several topoisomerase inhibitors have recently been shown to be active in preclinical systems. Topotecan (SK&F 104,864), a water-soluble camptothecin analog, is an inhibitor of topoisomerase I. Novobiocin is an inhibitor of topoisomerase II. Lonidamine depletes cellular adenosine 5'-triphosphate (ATP) and may impede energy-dependent DNA repair, MCF-7 human breast-cancer cells were treated in vitro with topotecan, novobiocin, and lonidamine alone, in paired combinations, and in combination with CDDP and melphalan. The three enzyme inhibitors alone and in combination did not increase tumor cell sensitivity to CDDP. However, the combinations of topotecan/novobiocin and lonidamine/novobiocin did enhance the cytotoxicity of melphalan. Mice bearing the FSaII fibrosarcoma were treated in vivo with topotecan, novobiocin, and lonidamine alone, in paired combinations, and in combination with CDDP, melphalan, BCNU, and cyclophosphamide. The combination of topotecan/novobiocin had the greatest impact on tumor cell sensitivity to each cytotoxic agent tested in both tumor cell-survival and tumor growth-delay assays. This sensitization was greatest at the highest concentrations of the cytotoxic agent tested. Combinations of topoisomerase I and topoisomerase II inhibitors may be useful as modulators of antitumor alkylating agents.
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PMID:Modulation of antitumor alkylating agents by novobiocin, topotecan, and lonidamine. 825 94


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